Differences between the left and right hemisphere of the brain have been observed in humans and rodents (Broca, 1861; Wernicke, 1881), including the hippocampal formation, a region of the brain that is necessary for some forms of learning and memory (Olton and Samuelson, 1976; deToledo-Morrell et al., 1988; Bernasconi-Guastalla et al., 1994; Tabibnia et al., 1994; Poe et al., 2000; Lister et al., 2006; Hanlon et al., 2005; Sommer et al., 2005; Moskal et al., 2006; Thompson et al., 2008; Klur et al., 2009). Although lateralization of the hippocampal formation has been studied in the adult, few have sought to directly examine the development of hippocampal lateralization (Moskal et al., 2006) and none have examined hippocampal lateralization in the embryo.
The objective of the study outlined in this dissertation was to characterize the development of hippocampal lateralization in the rat. To achieve this objective, a rat CNS microarray with 1,178 genes representing the majority of ontological categories within the rat genome (Kroes et al., 2006) was used to examine lateralized gene expression in the embryonic rat hippocampal formation: 14 genes were all more highly expressed in the
right hippocampus at E18 (Gross et al., 2008; Gross et al., 2010). Database for Annotation Visualization and Integrated Discovery (DAVID) and Gene Set Enrichment Analysis (GSEA) were also used to further investigate whether specific genes differentially expressed at E18 comprised pathways known to be important in the development of the hippocampal formation. Results demonstrated that genes related to structure, transcription and translation, cellular metabolism, glycolysis, and gap junction signaling were more highly expressed in the right hippocampus at E18. Expression of genes corresponding to proteins that comprise the gap junction signaling pathway were further examined using qRT-PCR. Results showed that alpha1a-tubulin, beta3-tubulin, and connexin43 were more highly expressed in the right hippocampus at E18. Using Western blot analysis, alpha1a-tubulin protein levels were also shown to be higher in the right hippocampus at E18. These results indicated that genes related to hippocampal growth and development were more highly expressed in the right hippocampus at E18, and furthermore they suggested that gap junctions may play a critical role in the development of hippocampal lateralization in the embryo.
To further characterize the lateralized development of the rat hippocampal formation, the effect of N-methyl-D-aspartate glutamate receptor (NMDAR) mediated synaptic activity lateralized gene expression in the hippocampal formation during early postnatal development in the rat. During normal development, the pattern of lateralized gene expression displays a right-to-left shift in preferential expression between P6 and P9 (Moskal et al., 2006). A reduction in NMDA receptor (NMDAR) mediated synaptic activity using the selective NMDAR antagonist CPP, altered this pattern of lateralized gene expression at P9 (Rahimi et al., 2006, Gross et al., 2007; Claiborne et al., 2010).
These data were then analyzed using Significance Analysis for Microarrays, DAVID, and GSEA analyses. The MAPK signaling pathway was enriched in the right hippocampal formation at P9 following CPP injections: these data were corroborated and extended using qRT-PCR. Expression of MAPK14 mRNA was not significantly different between the left and right hippocampal formation at postnatal day 6, nor was it greater in the right HF as compared to the left in saline treated rats at P9; however, MAPK14 mRNA was more highly expressed in the right hippocampal formation at P9 following a reduction in NMDAR activity between P6 and P9. c-Myc was more highly expressed in the right hippocampal formation at P6, and it was not differentially expressed during normal development or following saline control injections between P6 and P9. However, cMyc mRNA expression was significantly greater in the right hippocampal formation in CPP treated rats. These findings indicated that genes involved in the MAPK signaling pathway were upregulated in the right hippocampal formation during early postnatal development following a reduction in NMDAR-mediated synaptic activity.
The findings presented in this dissertation are both novel and important: they are the first to demonstrate that lateralized gene expression is present in the embryonic rat hippocampal formation. Furthermore, these findings are the first to show the effect of early experience on the development of hippocampal lateralization in the first postnatal week. The results support the idea that differential gene expression patterns in the hippocampus are likely developmentally regulated and play a key role in the formation and function of that region and that the gene expression patterns can be significantly influenced by factors that modulate synapse plasticity.